Effects of prostaglandin E2 on cells cultured from the rat organum vasculosum laminae terminalis and median preoptic nucleus.

The time course of the induction of enzymes responsible for the formation of prostaglandin E2 (PGE2) after an inflammatory insult, in relation to the concomitant febrile response, suggests that peripherally generated PGE2 is involved in the induction of the early phase of fever, while centrally produced PGE2 exerts pyrogenic capacities during the later stages of fever within the hypothalamic median preoptic nucleus (MnPO). The actions of peripherally derived PGE2 on the brain might occur at the level of the organum vasculosum laminae terminalis (OVLT), which lacks a tight blood-brain barrier and is implicated in fever, while the effects of PGE2 within the MnPO might interfere with glutamatergic neurotransmission within a recently characterized central efferent pathway for the activation of cold-defence reactions. Using the fura-2 ratio imaging technique we, therefore, measured changes of the intracellular Ca(2+)-concentration in primary neuroglial microcultures of rat OVLT and MnPO stimulated with PGE2 and/or glutamate. In cultures from the OVLT, as opposed to those derived from the MnPO, substantial numbers of neurons (8% of 385), astrocytes (19% of 645) and microglial cells (28% of 43) directly responded to PGE2 with a transient increase of intracellular Ca(2+). The most pronounced effect of PGE2 on cells from MnPO microcultures was its modulatory influence on the strength of glutamate-induced Ca(2+)-signals. In 72 out of 512 neurons and in 105 out of 715 astrocytes PGE2 significantly augmented glutamate-induced Ca(2+)-signals. About 30% of these neurons were GABAergic. These observations are in agreement with putative roles of peripheral PGE2 as a directly acting circulating agent at the level of the OVLT, and of central MnPO-intrinsic PGE2 as an enhancer of glutamatergic neurotransmission, which causes disinhibition of thermogenic heat production, a crucial component for the manifestation of fever. In microcultures from both brain sites investigated incubation with PGE2 significantly reduced the lipopolysaccharide-induced release of cytokines (tumor necrosis factor-α and interleukin-6) into the supernatant. PGE2, thus, seems to be involved in a negative feed-back loop to limit the strength of the brain inflammatory process and to play a dual role with pro- as well as anti-inflammatory properties.

Cytokine gene expression in murine fetal intestine: potential for extrathymic T cell development.

An increasing body of evidence suggests that certain T cell subsets mature extrathymically in the epithelium of the intestine. In the studies reported here, the authors have analysed cytokine/growth factor gene expression, recombinase-activating gene RAG-1 and RAG-2 gene expression and terminal deoxynucleotidyl transferase (TdT) gene expression in mouse fetal intestine and fetal thymus and liver, two known haematopoietic tissues. Stem cell factor (SCF) and interleukin 7 (IL-7) message was abundant in all three tissues during fetal development. IL-2 and IL-4 were not expressed in fetal gut but IL-4 was weakly detected in fetal liver and thymus. IL-9 and IL-13 mRNA was detected in all fetal tissues and IL-15 mRNA was abundant in fetal intestine but only weakly expressed in fetal liver and thymus. mRNA for SCF, IL-7, IL-13 and IL-15 was also detected in fibroblast-like cell lines derived from fetal intestine. RAG-1 and RAG-2 mRNA was detected in all three fetal tissues. TdT mRNA was not detected in fetal gut or liver but was weakly expressed in (fetal day) fd19-20 fetal thymus. Long-term (> 6 weeks) in vitro growth of IEL was achieved by coculturing intraepithelial lymphocytes (IEL) with IL-7-secreting fibroblasts in the presence of SCF and IL-2. The data show that the fetal mouse gut provides a suitable environment for lymphocyte development and receptor rearrangement, similar to fetal thymus and liver, even though expansion of intestinal IEL is delayed until 2-3 weeks after birth.

Impaired glucose homeostasis during postimplantation pregnancy in the mouse following acute exposure to ethanol, with particular reference to the uterus and embryo.

Dramatic changes in the levels of plasma glucose and lactate and liver glycogen were observed in mice, given an intraperitoneal injection of ethanol (3.5 g/kg body weight) on Day 9 of pregnancy, during the period of time (6 h) required to clear the drug from the circulatory system. These alterations were accompanied by significant changes in the rates of accumulation of some glycolytic and citric acid cycle intermediates in the uterus, including glucose-6-phosphate, fructose-6-phosphate, lactate, citrate, alpha-ketoglutarate, and succinate. Although the changes in some metabolic parameters were very transient, not all metabolites returned to control values by the time that the drug had been cleared from the maternal system. Alcohol also impaired the capacity of Day 9 mouse embryos to metabolize [14C]glucose under culture conditions in vitro and significantly increased the amount of the aldohexose accumulating in the fetal membrane fluid when administered on Day 14 of pregnancy. However, ethanol neither influenced the ratio of NADH to NAD+ in the uterus nor changed the glycolytic and respiratory activity of the uterine endometrium when coincubated with the tissue in vitro. The results indicate that glucose homeostasis is impaired in both the embryo and the maternal system of mice acutely exposed to alcohol during the teratogenically sensitive period of postimplantation pregnancy and support the thesis that this phenomenon may present an important mechanism underlying the embryo-toxic effects of alcohol consumed under "binge" drinking conditions during pregnancy. However, the results also suggest that the effects registered at the uterine level most likely involve stress reactions and acetate rather than primary actions of the drug on the organ.

Urinary corticotropin-releasing hormone immunoreactivity is elevated during human pregnancy.

Plasma corticotropin-releasing hormone immunoreactivity (CRH IR) rises with gestational age in women. In order to investigate the physiological changes of the hormone in pregnant women's urine, CRH IR was measured by radioimmunoassay in urine collected over a 24-hour period, a blood sample and a subsequent single collection of urine after the 24-hour collection (spot urine). Plasma CRH IR in pregnant subjects, 8682.8 +/- 2063.0 pg CRH IR/ml plasma (mean +/- SEM, n = 25), was significantly higher than that in the non-pregnant controls (7.2 +/- 1.6 pg/ml, n = 5; separate t = 4.21, p = 0.0003, d.f. = 24). Similarly, pregnant women had higher spot urine CRH IR - 54.6 +/- 15.5 pg/mumol creatinine (Cr) versus 5.0 +/- 0.5 pg/mumol Cr (separate t = 3.20, p = 0.0038, d.f. = 24.0) - and 24-hour urine CRH IR - 13.7 +/- 1.2 pg/mumol Cr compared with 7.7 +/- 0.8 pg/mumol Cr (separate t = 4.28, p = 0.003, d.f. = 24.4) than the non-pregnant cohort. The difference between urinary excretion of CRH IR as estimated by 24-hour urine (13.7 +/- 1.2 pg/mumol Cr) and spot urine (54.6 +/- 15.5 pg/mumol Cr) indicated that CRH IR in 24-hour urine may be degraded during storage. The weak associations between plasma and 24-hour urine CRH IR of pregnant women (correlation coefficient r = 0.34, p greater than 0.1), and total 24-hour urine and spot urine CRH IR (r = 0.25, p less than 0.1) further indicate CRH degradation. Plasma and spot urinary CRH IR, however, were strongly correlated (r = 0.80, p = 0.001). The total CRH IR excreted as estimated from the spot urine value (0.5 +/- 0.1 micrograms/day) compared with the total filtered load of CRH IR in the pregnant group (1306.9 +/- 324.6 micrograms/day) showed that 99.97% of the filtered CRH IR was reabsorbed or metabolized by the kidneys. Acidic gel chromatography of spot and 24-hour urine samples showed a CRH IR peak at CRH41 standard elution position (Kd = 0.5), indicating that the molecular form in urine is similar to the 41-residue standard. Pregnancy-induced hypertension correlated positively with plasma CRH IR (r = 0.62, p less than 0.001) and spot urine CRH IR (r = 0.46, p less than 0.01), and negatively with parity (r = -0.60, p less than 0.001). Plasma CRH IR and parity also negatively correlated (r = -0.41, p less than 0.05).(ABSTRACT TRUNCATED AT 400 WORDS)

The role of acetate in alcohol-induced alterations of uterine glucose metabolism in the mouse during pregnancy.

The acute exposure of mice to ethanol during post-implantation pregnancy has been reported to cause alterations in the levels of several glycolytic intermediates in the uterus, suggesting a possible indirect mechanism of alcohol embryo-toxicity. The present study was undertaken to assess whether the ethanol metabolite, acetate is implicated in this phenomenon. Blood and uterine alcohol concentrations in day 9--pregnant Quackenbush Swiss mice were maximal 15 minutes after the intraperitoneal injection of ethanol (3.5 g/kg body weight), and fell to almost negligible levels 6 hours later. In response to this treatment, the levels of blood and uterine acetate increased, liver glycogen decreased, plasma glucose increased, and uterine glucose, glucose-6-phosphate (G-6-P), fructose-6-phosphate (F-6-P), and citrate increased. When acetate was administered to pregnant mice in amounts approximating those generated by exposure to alcohol, the levels of uterine F-6-P and citrate increased while other metabolic parameters remained unaffected. The administration of 4-methylpyrazole to mice subsequently treated with alcohol produced conditions of alcohol exposure in the absence of ethanol-derived acetate and depressed the ethanol-induced rise in uterine G-6-P and citrate. The results support the notion that acetate contributes to the alcohol-induced alterations in metabolism, at least as far as the regulation of uterine citrate and hexose monophosphates are concerned. This, together with stress responses induced by exposure to the acute dose of alcohol, may present mechanisms underlying the fetal alcohol syndrome associated in particular with "binge" drinking.